This preclinical project combines imaging technology development with biomarker discovery for functional image guidance during resection of meduloblastoma (MB), the most common pediatric brain tumor. A novel imaging technology, the dual-axis confocal (DAC) microscope, will be developed to image fluorescent probes that target MB biomarkers. First, a tabletop DAC will be used, along with targeted probes, to image a spontaneous mouse tumor model of MB. A quantitative ratiometric-imaging method will be employed to improve tumor-to-normal image contrast, which will also be maximized by optimizing parameters such as imaging depth, probe concentration, and the procedure for rinse removal of unbound probe. A miniature DAC, which incorporates a 2-mm diameter gradient index needle lens, will be developed to guide MB debulking surgeries in mice. Improved resection will be demonstrated by quantifying residual tumor mass, time-to-relapse, and by monitoring overall survival. These technologies and methods will enable more complete and accurate resection of MB in the brain, and will thereby improve patient outcomes, by allowing surgeons to visualize tumor margins with cellular accuracy and molecular specificity.